Anthramycin is an antitumor antibiotic produced by Streptomyces refuineus. The mode of action of this anticancer agent is to selectively inhibit both RNA and DNA polymerases by binding to the DNA template. It is strongly suspected that anthramycin binds to DNA through formation of a covalent linkage. However, the point at which this linkage between anthramycin and DNA occurs is unknown. As a consequence of biosynthetic feeding experiments with the anthramycin producing strain of S. refuineus we have on hand a variety of specifically tritiated and carbon-14 labelled anthramycin molecules, and in addition the capability to prepare specifically labelled carbon-13 anthramycin molecules. We propose to use these specifically labelled anthramycin molecules to determine the structure of the anthramycin-DNA complex. Firstly it is proposed to establish which, if any, of the non-exchangeable hydrogen or carbon atoms of anthramycin are lost during formation of the anthramycin-DNA complex. This will be achieved by a comparison of the tritium to carbon-14 ratios of the specifically labelled anthramycin molecules with that of the anthramycin-DNA complex formed after incubation of the DNA with these same specifically labelled anthramycin molecules. Secondly it is proposed to further establish the structure of this complex by a study using carbon-13 NMR. In this study anthramycin-DNA molecules specifically enriched with C-13 will be used to produce the anthramycin-DNA complex. The C-13 NMR chemical shifts and spin lattice relaxation times of those carbons of the complex enriched with C-13 will then be used to establish criteria upon which to build a model for the anthramycin-DNA complex. Since it is essential that the biosynthetic labelling pattern for anthramycin is unequivocally known, a series of biosynthetic studies utilizing carbon-13 and deuterium will be conducted to establish this pattern. BIBLIOGRAPHIC REFERENCES: L. Hurley, M. Zmijewski and C. J. Chang, Biosynthesis of Antrhamycin. Determination of the labelling pattern by the Use of Radioactive and Stable Isotope Techniques. J. Am. Chem. Soc., 97, 4372 (1975).